The IGF ligands, IGF-I and IGF-II, and the IGF type I receptor (IGF-IR) have demonstrated roles in growth of mammary epithelium during normal development. Moreover, the IGFs and IGF-IR promote proliferation of breast cancer cells and are implicated in incidence and growth of breast cancers. Our previous studies demonstrated that IGF-I and IGF-II are distinctly expressed during pubertal and pregnancy-induced development of the mammary epithelium. During the previous funding period, we demonstrated that IGF-I regulates cell cycle progression in mammary epithelial cells and showed unique roles for IGF-I expressed in epithelial and stromal compartments of the developing mammary gland. We further elucidated how patterning of IGF-II expression is regulated in the mammary epithelium. Recent data suggest that the IGF ligands can signal through heterodimers of the IGF-IR and the insulin receptor (IR) in addition to homodimers of the IGF-IR. Moreover, a splice variant of the IR known as the IR-A isoform, has high specificity for IGF-II but not IGF-I. Our preliminary data show that mammary specific deletion of the IR during alveolar differentiation results in significant disruption of alveolar development and lactation. Furthermore, analysis of receptor expression suggests that the ratio of the two IR isoforms varies in mammary epithelial cells during pregnancy and lactation and that a significant portion of the IGF-IR exists as a hybrid receptor with the IR in mammary epithelial cells. Thus, the goal of the experiments proposed in this application is to test the hypothesis that the different IGF signaling receptors have distinct functions in alveolar differentiation and lactation. To test this hypothesis, we propose both in vitro experiments and in vivo transgenic studies to activate or disrupt signaling through the IRs, IGF-IR and hybrid receptors in mammary epithelial cells. Public Health Relevance Statement: The results of the proposed experiments will elucidate IGF receptor-specific actions and downstream signaling pathways in proliferation, survival and differentiation of normal mammary epithelial cells. These studies will also have important implications for the roles of these receptors and pathways in breast cancers.